NS5B

About: NS5B is a research topic. Over the lifetime, 1314 publications have been published within this topic receiving 59534 citations.

RNA aptamer-mediated interference of HCV replication by targeting the CRE-5BSL3.2 domain

Abstract: The RNA genome of hepatitis C virus (HCV)contains multiple conserved structural RNA domains thatplay key roles in essential viral processes. A conservedstructural component within the 3¢ end of the region codingfor viral RNA-dependent RNA polymerase (NS5B) has beencharacterized as a functional cis-acting replication element(CRE). This study reports the ability of two RNA aptamers,P-58 and P-78, to interfere with HCV replication by tar-geting the essential 5BSL3.2 domain within this CRE.Structure-probing assays showed the binding of the apta-mers to the CRE results in a structural reorganization of theapical portion of the 5BSL3.2 stem-loop domain. Thisinterfered with the binding of the NS5B protein to the CREand induced a significant reduction in HCV replication(»50%) in an autonomous subgenomic HCV replicationsystem. These results highlight the potential of this CRE as atarget for the development of anti-HCV therapies andunderscore the potential of antiviral agents based on RNAaptamer molecules.Keywords: CRE, HCV genome, RNA aptamers, RNA structurefunction.

Allosteric mechanism of cyclopropylindolobenzazepine inhibitors for HCV NS5B RdRp via dynamic correlation network analysis

Abstract: HCV RNA dependent RNA polymerase (RdRp) nonstructural protein 5B (NS5B) is a major target against hepatitis C virus (HCV) for antiviral therapy. Recently discovered cyclopropylindolobenzazepine derivatives have been considered as the most potent for their ability to bind the thumb site 1 domain and allosterically inhibit HCV NS5B RdRp activity. However, the allosteric mechanism for these derivatives has not been clarified at the molecular level. In this study, fluctuation correlation networks were constructed based on all-atom molecular dynamics simulations to elucidate the allosteric mechanism. The fluctuation correlation networks between free and M2 bound NS5B are significantly different. Information can better transfer from the allosteric site to the catalytic site for bound NS5B than for free NS5B. Thus, the hypothesis of "binding induced allosteric regulation" is proposed to link the enzyme activation and inhibitor binding and then confirmed by the mutant network. Finally, one possible allosteric pathway was identified with the shortest path and evaluated by the perturbation of the network. These methods will be helpful to identify the allosteric pathway of other proteins and to design new drugs targeting the pathway.

Site-specific cleavage of HCV genomic RNA and its cloned core and NS5B genes by DNAzyme.

Abstract: Background and Aims: The 9600 nt hepatitis C virus (HCV) genomic RNA has only one internal ribosome entry site (IRES) for translation to a single polyprotein. In search of nucleic acid-based antiviral agents, two 10-23 DNAzymes were designed to cleave the RNA in IRES and RNA dependent RNA polymerase (RDRP/NS5B) regions to prevent translation and replication of HCV RNA. Methods: In vitro cleavage of HCV RNA by IRES specific DNAzyme, CDz and NS5B specific DNAzyme, NDz was carried out using HCV genomic RNA and in vitro synthesized runoff transcripts of core and NS5B genes. Cleavage of core and NS5B mRNAs by DNAzyme (Dz) in HepG2 cells was assessed by reverse transcription polymerase chain reaction (RT-PCR) using RNA from cells co-transfected with cloned core or NS5B gene and its respective DNAzyme. Suppression of core or NS5B protein expression due to mRNA cleavage by Dz in co-transfected cells was determined by Western blot analysis and fluorescence intensity of fluorescent-tagged expressed protein. Reduction of NS5B protein activity in NDz co-transfected cells was determined by enzymatic assays. Results: The designed CDz and NDz cleaved HCV genomic RNA and their respective in vitro generated transcripts. Both mRNA and protein expressions of core or NS5B from their cloned genes reduced substantially when co-transfected with respective Dz. Reduction of RDRP expression by NDz was accompanied with its reduced enzyme activity. Increased RNA cleavage, inhibition of protein expression, and reduction of RDRP activity were observed on increasing Dz concentration. Conclusion: Core and NS5B targeted DNAzymes can be used in controlling the replication of HCV RNA.

Cyclophilin A as a New Therapeutic Target for Hepatitis C Virus-induced Hepatocellular Carcinoma.

Abstract: Hepatocellular carcinoma (HCC) related to hepatitis B virus (HBV) and hepatitis C virus (HCV) infections is thought to account for more than 80% of primary liver cancers. Both HBV and HCV can establish chronic liver inflammatory infections, altering hepatocyte and liver physiology with potential liver disease progression and HCC development. Cyclophilin A (CypA) has been identified as an essential host factor for the HCV replication by physically interacting with the HCV non structural protein NS5A that in turn interacts with RNA-dependent RNA polymerase NS5B. CypA, a cytosolic binding protein of the immunosuppressive drug cyclosporine A, is overexpressed in many cancer types and often associated with malignant transformation. Therefore, CypA can be a good target for molecular cancer therapy. Because of antiviral activity, the CypA inhibitors have been tested for the treatment of chronic hepatitis C. Nonimmunosuppressive Cyp inhibitors such as NIM811, SCY-635, and Alisporivir have attracted more interests for appropriating CypA for antiviral chemotherapeutic target on HCV infection. This review describes CypA inhibitors as a potential HCC treatment tool that is contrived by their obstructing chronic HCV infection and summarizes roles of CypA in cancer development.